As the most abundant liver-specific microRNA (miRNA), miR-122 has been extensively studied for its role in the regulation of lipid metabolism, hepatocarcinogenesis and hepatitis C virus (HCV) replication, but little is known regarding its role in the replication of Hepatitis B virus (HBV), a highly prevalent hepatotropic virus that can cause life-threatening complications. In this study we examined the effects of antisense inhibition of miR-122 and transfection of a miR-122 mimic on HBV expression in hepatoma cells. The over-expression of miR-122 inhibited HBV expression, whereas the depletion of endogenous miR-122 resulted in increased production of HBV in transfected cells. We further found that the down-regulation of Heme oxygenase-1 (HO-1) by miR-122 plays a negative role in the miR-122-mediated inhibition of viral expression. Our study demonstrates the anti-HBV activity of miR-122, suggesting that therapies that increase miR-122 and HO-1 may be an effective strategy to limit HBV replication. (C) 2010 Elsevier Inc. All rights reserved.